Deregulation of KSHV latency conformation by ER-stress and caspase-dependent RAD21-cleavage

Kaposi’s sarcoma (KS)-associated herpesvirus (KSHV) is a human gammaherpesvirus recognized as the principal causative agent of KS and primary effusion lymphoma (PEL). KSHV establishes persistent latent infection in B lymphocytes where viral gene expression is restricted, in part, by a cohesin-dependent chromosome conformation. Here, we show that endoplasmic reticulum (ER) stress induces a rapid, caspase-dependent cleavage of cohesin subunit RAD21. ER stress-induced cleavage of RAD21 correlated with a rapid and strong viral lytic transcriptional activation. This effect was observed in several KSHV positive PEL cells, but not in other B-cells or non-B-cell models of KSHV latency. The cleaved-RAD21 does not dissociate from viral genomes, nor disassemble from other components of the cohesin complex. However, RAD21 cleavage correlated with the disruption of the latency genome conformation as revealed by chromosome conformation capture (3C). Ectopic expression of C-terminal RAD21 cleaved form could partially induce KSHV lytic genes transcription in BCBLI cells, suggesting that ER-stress induced RAD21 cleavage was sufficient to induce KSHV reactivation from latency in PEL cells. Taken together our results reveal a novel aspect for control and maintenance of KSHV genome latency conformation mediated by stress-induced RAD21 cleavage. Our studies also suggest that RAD21 cleavage may be a general regulatory mechanism for rapid alteration of cellular chromosome conformation and cohesin-dependent transcription regulation.卡波西肉瘤（Kaposi’s sarcoma, KS）相关疱疹病毒（Kaposi’s sarcoma-associated herpesvirus, KSHV）是一种人类γ疱疹病毒，被认定为卡波西肉瘤与原发性渗出性淋巴瘤（primary effusion lymphoma, PEL）的主要致病病原体。KSHV可在B淋巴细胞中建立持续性潜伏感染，该过程中病毒基因表达受到严格限制，其中部分调控机制依赖于黏连蛋白（cohesin）介导的染色体构象调控。本研究证实，内质网（endoplasmic reticulum, ER）应激可快速诱导黏连蛋白亚基RAD21发生半胱天冬酶依赖的蛋白裂解。内质网应激介导的RAD21裂解，与病毒快速且强效的裂解型转录激活呈显著相关性。该效应在多种KSHV阳性的原发性渗出性淋巴瘤细胞中均有观测到，但在其他B细胞或非B细胞来源的KSHV潜伏感染模型中并未出现。裂解后的RAD21既不会从病毒基因组上脱离，也不会与黏连蛋白复合物的其他组分发生解聚。然而，通过染色体构象捕获技术（chromosome conformation capture, 3C）分析可见，RAD21的裂解与潜伏感染的病毒基因组构象破坏密切相关。异位表达RAD21的C端裂解片段，可在BCBLI细胞中部分诱导KSHV裂解基因的转录，这表明内质网应激诱导的RAD21裂解足以触发原发性渗出性淋巴瘤细胞中KSHV从潜伏状态向裂解状态的重新激活。综上，本研究结果揭示了应激诱导的RAD21裂解在调控与维持KSHV基因组潜伏构象中的全新作用途径。同时，本研究还提示RAD21裂解可能是一种通用的细胞调控机制，可快速改变细胞染色体构象并调控黏连蛋白依赖的转录过程。